Discovery of biomarkers in the psoriasis through machine learning and dynamic immune infiltration in three types of skin lesions.

Introduction: Psoriasis is a chronic skin disease characterized by unique scaling plaques. However, during the acute phase, psoriatic lesions exhibit eczematous changes, making them difficult to distinguish from atopic dermatitis, which poses challenges for the selection of biological agents. This study aimed to identify potential diagnostic genes in psoriatic lesions and investigate their clinical significance. Methods: GSE182740 datasets from the GEO database were analyzed for differential analysis; machine learning algorithms (SVM-RFE and LASSO regression models) are used to screen for diagnostic markers; CIBERSORTx is used to determine the dynamic changes of 22 different immune cell components in normal skin lesions, psoriatic non-lesional skin, and psoriatic lesional skin, as well as the expression of the diagnostic genes in 10 major immune cells, and real-time quantitative polymerase chain reaction (RT-qPCR) and immunohistochemistry are used to validate results. Results: We obtained 580 differentially expressed genes (DEGs) in the skin lesion and non-lesion of psoriasis patients, 813 DEGs in mixed patients between non-lesions and lesions, and 96 DEGs in the skin lesion and non-lesion of atopic dermatitis, respectively. Then 144 specific DEGs in psoriasis via a Veen diagram were identified. Ultimately, UGGT1, CCNE1, MMP9 and ARHGEF28 are identified for potential diagnostic genes from these 144 specific DEGs. The value of the selected diagnostic genes was verified by receiver operating characteristic (ROC) curves with expanded samples. The the area under the ROC curve (AUC) exceeded 0.7 for the four diagnosis genes. RT-qPCR results showed that compared to normal human epidermis, the expression of UGGT1, CCNE1, and MMP9 was significantly increased in patients with psoriasis, while ARHGEF28 expression was significantly decreased. Notably, the results of CIBERSORTx showed that CCNE1 was highly expressed in CD4+ T cells and neutrophils, ARHGEF28 was also expressed in mast cells. Additionally, CCNE1 was strongly correlated with IL-17/CXCL8/9/10 and CCL20. Immunohistochemical results showed increased nuclear expression of CCNE1 in psoriatic epidermal cells relative to normal. Conclusion: Based on the performance of the four genes in ROC curves and their expression in immune cells from patients with psoriasis, we suggest that CCNE1 possess higher diagnostic value.

Multimodal Therapy Approaches for NUT Carcinoma by Dual Combination of Oncolytic Virus Talimogene Laherparepvec with Small Molecule Inhibitors.

NUT (nuclear-protein-in-testis) carcinoma (NC) is a highly aggressive tumor disease. Given that current treatment regimens offer a median survival of six months only, it is likely that this type of tumor requires an extended multimodal treatment approach to improve prognosis. In an earlier case report, we could show that an oncolytic herpes simplex virus (T-VEC) is functional in NC patients. To identify further combination partners for T-VEC, we have investigated the anti-tumoral effects of T-VEC and five different small molecule inhibitors (SMIs) alone and in combination in human NC cell lines. Dual combinations were found to result in higher rates of tumor cell reductions when compared to the respective monotherapy as demonstrated by viability assays and real-time tumor cell growth monitoring. Interestingly, we found that the combination of T-VEC with SMIs resulted in both stronger and earlier reductions in the expression of c-Myc, a main driver of NC cell proliferation, when compared to T-VEC monotherapy. These results indicate the great potential of combinatorial therapies using oncolytic viruses and SMIs to control the highly aggressive behavior of NC cancers and probably will pave the way for innovative multimodal clinical studies in the near future.

AURKB promotes colorectal cancer progression by triggering the phosphorylation of histone H3 at serine 10 to activate CCNE1 expression.

Aurora kinase B (AURKB) initiates the phosphorylation of serine 10 on histone H3 (pH3S10), a crucial process for chromosome condensation and cytokinesis in mammalian mitosis. Nonetheless, the precise mechanisms through which AURKB regulates the cell cycle and contributes to tumorigenesis as an oncogenic factor in colorectal cancer (CRC) remain unclear. Here, we report that AURKB was highly expressed and positively correlated with Ki-67 expression in CRC. The abundant expression of AURKB promotes the growth of CRC cells and xenograft tumors in animal model. AURKB knockdown substantially suppressed CRC proliferation and triggered cell cycle arrest in G2/M phase. Interestingly, cyclin E1 (CCNE1) was discovered as a direct downstream target of AURKB and functioned synergistically with AURKB to promote CRC cell proliferation. Mechanically, AURKB activated CCNE1 expression by triggering pH3S10 in the promoter region of CCNE1. Furthermore, it was showed that the inhibitor specific for AURKB (AZD1152) can suppress CCNE1 expression in CRC cells and inhibit tumor cell growth. To conclude, this research demonstrates that AURKB accelerated the tumorigenesis of CRC through its potential to epigenetically activate CCNE1 expression, suggesting AURKB as a promising therapeutic target in CRC.

Phospho-mimetic CD3ε variants prevent TCR and CAR signaling.

Introduction: Antigen binding to the T cell antigen receptor (TCR) leads to the phosphorylation of the immunoreceptor tyrosine-based activation motifs (ITAMs) of the CD3 complex, and thereby to T cell activation. The CD3ε subunit plays a unique role in TCR activation by recruiting the kinase LCK and the adaptor protein NCK prior to ITAM phosphorylation. Here, we aimed to investigate how phosphorylation of the individual CD3ε ITAM tyrosines impacts the CD3ε signalosome. Methods: We mimicked irreversible tyrosine phosphorylation by substituting glutamic acid for the tyrosine residues in the CD3ε ITAM. Results: Integrating CD3ε phospho-mimetic variants into the complete TCR-CD3 complex resulted in reduced TCR signal transduction, which was partially compensated by the involvement of the other TCR-CD3 ITAMs. By using novel CD3ε phospho-mimetic Chimeric Antigen Receptor (CAR) variants, we avoided any compensatory effects of other ITAMs in the TCR-CD3 complex. We demonstrated that irreversible CD3ε phosphorylation prevented signal transduction upon CAR engagement. Mechanistically, we demonstrated that glutamic acid substitution at the N-terminal tyrosine residue of the CD3ε ITAM (Y39E) significantly reduces NCK binding to the TCR. In contrast, mutation at the C-terminal tyrosine of the CD3ε ITAM (Y50E) abolished LCK recruitment to the TCR, while increasing NCK binding. Double mutation at the C- and N-terminal tyrosines (Y39/50E) allowed ZAP70 to bind, but reduced the interaction with LCK and NCK. Conclusions: The data demonstrate that the dynamic phosphorylation of the CD3ε ITAM tyrosines is essential for CD3ε to orchestrate optimal TCR and CAR signaling and highlights the key role of CD3ε signalosome to tune signal transduction.

UHRF2 accumulates in early G1-phase after serum stimulation or mitotic exit to extend G1 and total cell cycle length.

Ubiquitin like with PHD and ring finger domains 2 (UHRF2) regulates the cell cycle and epigenetics as a multi-domain protein sharing homology with UHRF1. UHRF1 functions with DNMT1 to coordinate daughter strand methylation during DNA replication, but UHRF2 can't perform this function, and its roles during cell cycle progression are not well defined. UHRF2 role as an oncogene vs. tumor suppressor differs in distinct cell types. UHRF2 interacts with E2F1 to control Cyclin E1 (CCNE1) transcription. UHRF2 also functions in a reciprocal loop with Cyclin E/CDK2 during G1, first as a direct target of CDK2 phosphorylation, but also as an E3-ligase with direct activity toward both Cyclin E and Cyclin D. In this study, we demonstrate that UHRF2 is expressed in early G1 following either serum stimulation out of quiescence or in cells transiting directly out of M-phase, where UHRF2 protein is lost. Further, UHRF2 depletion in G2/M is reversed with a CDK1 specific inhibitor. UHRF2 controls expression levels of cyclins and CDK inhibitors and controls its own transcription in a negative-feedback loop. Deletion of UHRF2 using CRISPR/Cas9 caused a delay in passage through each cell cycle phase. UHRF2 loss culminated in elevated levels of cyclins but also the CDK inhibitor p27KIP1, which regulates G1 passage, to reduce retinoblastoma phosphorylation and increase the amount of time required to reach G1/S passage. Our data indicate that UHRF2 is a central regulator of cell-cycle pacing through its complex regulation of cell cycle gene expression and protein stability.

Modulation of GPR133 (ADGRD1) signaling by its intracellular interaction partner extended synaptotagmin 1.

GPR133 (ADGRD1) is an adhesion G-protein-coupled receptor that signals through Gαs/cyclic AMP (cAMP) and is required for the growth of glioblastoma (GBM), an aggressive brain malignancy. The regulation of GPR133 signaling is incompletely understood. Here, we use proximity biotinylation proteomics to identify ESYT1, a Ca2+-dependent mediator of endoplasmic reticulum-plasma membrane bridge formation, as an intracellular interactor of GPR133. ESYT1 knockdown or knockout increases GPR133 signaling, while its overexpression has the opposite effect, without altering GPR133 levels in the plasma membrane. The GPR133-ESYT1 interaction requires the Ca2+-sensing C2C domain of ESYT1. Thapsigargin-mediated increases in cytosolic Ca2+ relieve signaling-suppressive effects of ESYT1 by promoting ESYT1-GPR133 dissociation. ESYT1 knockdown or knockout in GBM slows tumor growth, suggesting tumorigenic functions of ESYT1. Our findings demonstrate a mechanism for the modulation of GPR133 signaling by increased cytosolic Ca2+, which reduces the signaling-suppressive interaction between GPR133 and ESYT1 to raise cAMP levels.

The MYCN oncoprotein is an RNA-binding accessory factor of the nuclear exosome targeting complex.

The MYCN oncoprotein binds active promoters in a heterodimer with its partner protein MAX. MYCN also interacts with the nuclear exosome, a 3'-5' exoribonuclease complex, suggesting a function in RNA metabolism. Here, we show that MYCN forms stable high-molecular-weight complexes with the exosome and multiple RNA-binding proteins. MYCN binds RNA in vitro and in cells via a conserved sequence termed MYCBoxI. In cells, MYCN associates with thousands of intronic transcripts together with the ZCCHC8 subunit of the nuclear exosome targeting complex and enhances their processing. Perturbing exosome function results in global re-localization of MYCN from promoters to intronic RNAs. On chromatin, MYCN is then replaced by the MNT(MXD6) repressor protein, inhibiting MYCN-dependent transcription. RNA-binding-deficient alleles show that RNA-binding limits MYCN's ability to activate cell growth-related genes but is required for MYCN's ability to promote progression through S phase and enhance the stress resilience of neuroblastoma cells.

AGR2-mediated unconventional secretion of 14-3-3ε and α-actinin-4, responsive to ER stress and autophagy, drives chemotaxis in canine mammary tumor cells.

BACKGROUND: Canine mammary tumors (CMTs) in intact female dogs provide a natural model for investigating metastatic human cancers. Our prior research identified elevated expression of Anterior Gradient 2 (AGR2), a protein disulfide isomerase (PDI) primarily found in the endoplasmic reticulum (ER), in CMT tissues, highly associated with CMT progression. We further demonstrated that increased AGR2 expression actively influences the extracellular microenvironment, promoting chemotaxis in CMT cells. Unraveling the underlying mechanisms is crucial for assessing the potential of therapeutically targeting AGR2 as a strategy to inhibit a pro-metastatic microenvironment and impede tumor metastasis. METHODS: To identify the AGR2-modulated secretome, we employed proteomics analysis of the conditioned media (CM) from two CMT cell lines ectopically expressing AGR2, compared with corresponding vector-expressing controls. AGR2-regulated release of 14-3-3ε (gene: YWHAE) and α-actinin 4 (gene: ACTN4) was validated through ectopic expression, knockdown, and knockout of the AGR2 gene in CMT cells. Extracellular vesicles derived from CMT cells were isolated using either differential ultracentrifugation or size exclusion chromatography. The roles of 14-3-3ε and α-actinin 4 in the chemotaxis driven by the AGR2-modulated CM were investigated through gene knockdown, antibody-mediated interference, and recombinant protein supplement. Furthermore, the clinical relevance of the release of 14-3-3ε and α-actinin 4 was assessed using CMT tissue-immersed saline and sera from CMT-afflicted dogs. RESULTS: Proteomics analysis of the AGR2-modulated secretome revealed increased abundance in 14-3-3ε and α-actinin 4. Ectopic expression of AGR2 significantly increased the release of 14-3-3ε and α-actinin 4 in the CM. Conversely, knockdown or knockout of AGR2 expression remarkably reduced their release. Silencing 14-3-3ε or α-actinin 4 expression diminished the chemotaxis driven by AGR2-modulated CM. Furthermore, AGR2 controls the release of 14-3-3ε and α-actinin 4 primarily via non-vesicular routes, responding to the endoplasmic reticulum (ER) stress and autophagy activation. Knockout of AGR2 resulted in increased α-actinin 4 accumulation and impaired 14-3-3ε translocation in autophagosomes. Depletion of extracellular 14-3-3ε or α-actinin 4 reduced the chemotaxis driven by AGR2-modulated CM, whereas supplement with recombinant 14-3-3ε in the CM enhanced the CM-driven chemotaxis. Notably, elevated levels of 14-3-3ε or α-actinin 4 were observed in CMT tissue-immersed saline compared with paired non-tumor samples and in the sera of CMT dogs compared with healthy dogs. CONCLUSION: This study elucidates AGR2's pivotal role in orchestrating unconventional secretion of 14-3-3ε and α-actinin 4 from CMT cells, thereby contributing to paracrine-mediated chemotaxis. The insight into the intricate interplay between AGR2-involved ER stress, autophagy, and unconventional secretion provides a foundation for refining strategies aimed at impeding metastasis in both canine mammary tumors and potentially human cancers.

Molecular Landscape and Clinical Implication of CCNE1-amplified Esophagogastric Cancer.

Cyclin E overexpression as a result of CCNE1 amplification is a critical driver of genomic instability in gastric cancer, but its clinical implication is largely unknown. Thus, we integrated genomic, transcriptomic, and immune profiling analysis of 7,083 esophagogastric tumors and investigated the impact of CCNE1 amplification on molecular features and treatment outcomes. We identified CCNE1 amplification in 6.2% of esophageal adenocarcinoma samples, 7.0% of esophagogastric junction carcinoma, 4.2% of gastric adenocarcinoma samples, and 0.8% of esophageal squamous cell carcinoma. Metastatic sites such as lymph node and liver showed an increased frequency of CCNE1 amplification relative to primary tumors. Consistent with a chromosomal instability phenotype, CCNE1 amplification was associated with decreased CDH1 mutation and increased TP53 mutation and ERBB2 amplification. We observed no differences in immune biomarkers such as PD-L1 expression and tumor mutational burden comparing CCNE1-amplified and nonamplified tumors, although CCNE1 amplification was associated with changes in immune populations such as decreased B cells and increased M1 macrophages from transcriptional analysis. Real-world survival analysis demonstrated that patients with CCNE1-amplified gastric cancer had worse survival after trastuzumab for HER2-positive tumors, but better survival after immunotherapy. These data suggest that CCNE1-amplified gastric cancer has a distinct molecular and immune profile with important therapeutic implications, and therefore further investigation of CCNE1 amplification as a predictive biomarker is warranted. SIGNIFICANCE: Advanced gastric cancer has a relatively dismal outcome with a 5-year overall survival of less than 10%. Furthermore, while comprehensive molecular analyses have established molecular subtypes within gastric cancers, biomarkers of clinical relevance in this cancer type are lacking. Overall, this study demonstrates that CCNE1 amplification is associated with a distinct molecular profile in gastric cancer and may impact response to therapy, including targeted therapy and/or immunotherapy.

KLF4 suppresses anticancer effects of brusatol via transcriptional upregulating NCK2 expression in melanoma.

Brusatol (Bru), a main extract from traditional Chinese medicine Brucea javanica, has been reported to exist antitumor effect in many tumors including melanoma. However, the underlying mechanism in its anti-melanoma effect still need further exploration. Here, we reported that the protein expression of KLF4 in melanoma cells were significantly downregulated in response to brusatol treatment. Overexpression of KLF4 suppressed brusatol-induced melanoma cell apoptosis; while knockdown of KLF4 enhanced antitumor effects of brusatol on melanoma cells not only in vitro but also in vivo. Further studies on the mechanism revealed that KLF4 bound to the promoter of NCK2 directly and facilitated NCK2 transcription, which suppressed the antitumor effect of brusatol on melanoma. Furthermore, our findings showed that miR-150-3p was dramatically upregulated under brusatol treatment which resulted in the downregulation of KLF4. Our results suggested that the miR-150-3p/KLF4/NCK2 axis might play an important role in the antitumour effects of brusatol in melanoma.

Research progress on oncoprotein hepatitis B X­interacting protein (Review).

Hepatitis B X­interacting protein (HBXIP) is a membrane protein located on the lysosomal surface and encoded by the Lamtor gene. It is expressed by a wide range of tumor types, including breast cancer, esophageal squamous cell carcinoma and hepatocellular carcinoma, and its expression is associated with certain clinicopathological characteristics. In the past decade, research on the oncogenic mechanisms of HBXIP has increased and the function of HBXIP in normal cells has been gradually elucidated. In the present review, the following was discussed: The normal physiological role of the HBXIP carcinogenic mechanism; the clinical significance of high levels of HBXIP expression in different tumors; HBXIP regulation of transcription, post­transcription and post­translation processes in tumors; the role of HBXIP in improving the antioxidant capacity of tumor cells; the inhibition of ferroptosis of tumor cells and regulating the metabolic reprogramming of tumor cells; and the role of HBXIP in promoting the malignant progression of tumors. In conclusion, the present review summarized the existing knowledge of HBXIP, established its carcinogenic mechanism and discussed future related research on HBXIP.

RAD52-dependent mitotic DNA synthesis is required for genome stability in Cyclin E1-overexpressing cells.

Overexpression of Cyclin E1 perturbs DNA replication, resulting in DNA lesions and genomic instability. Consequently, Cyclin E1-overexpressing cancer cells increasingly rely on DNA repair, including RAD52-mediated break-induced replication during interphase. We show that not all DNA lesions induced by Cyclin E1 overexpression are resolved during interphase. While DNA lesions upon Cyclin E1 overexpression are induced in S phase, a significant fraction of these lesions is transmitted into mitosis. Cyclin E1 overexpression triggers mitotic DNA synthesis (MiDAS) in a RAD52-dependent fashion. Chemical or genetic inactivation of MiDAS enhances mitotic aberrations and persistent DNA damage. Mitosis-specific degradation of RAD52 prevents Cyclin E1-induced MiDAS and reduces the viability of Cyclin E1-overexpressing cells, underscoring the relevance of RAD52 during mitosis to maintain genomic integrity. Finally, analysis of breast cancer samples reveals a positive correlation between Cyclin E1 amplification and RAD52 expression. These findings demonstrate the importance of suppressing mitotic defects in Cyclin E1-overexpressing cells through RAD52.

Whole-transcriptome profiling reveals potential biomarkers for the reversal of thymic epithelial cell senescence by umbilical cord mesenchymal stem cells.

BACKGROUND: Reduced numbers and dysfunction of thymic epithelial cells (TECs) are important factors of thymic degeneration. Previous studies have found that umbilical cord mesenchymal stem cells (UCMSCs) reverse the structure and function of the senescent thymus in vivo. However, the transcriptomic regulation mechanism is unclear. METHODS: TECs were cultured with H2O2 for 72 hours to induce senescence. UCMSCs were cocultured with senescent TECs for 48 hours to detect SA-ß-gal, P16 and Ki67. The cocultured TECs were collected for lncRNA, mRNA and miRNA sequencing to establish a competitive endogenous regulatory network (ceRNA). And RT-qPCR, immunofluorescence staining, and western blot were used to identified key genes. RESULTS: Our results showed that H2O2 induced TEC aging and that UCMSCs reversed these changes. Compared with those in aged TECs, 2260 DE mRNAs, 1033 DE lncRNAs and 67 DE miRNAs were differentially expressed, and these changes were reversed by coculturing the cells with UCMSCs. Differential mRNA enrichment analysis of ceRNA regulation revealed that the PI3K-AKT pathway was a significant signaling pathway. UCMSC coculture upregulated VEGFA, which is the upstream factor of the PI3K-AKT signaling pathway, and the expression of the key proteins PI3K and AKT. Thus, the expression of the cell cycle suppressor P27, which is downstream of the PI3K-AKT signaling pathway, was downregulated, while the expression of the cell cycle regulators CDK2 and CCNE was upregulated. CONCLUSION: UCMSC coculture upregulated the expression of VEGFA, activated the PI3K-AKT signaling pathway, increased the expression of CDK2 and CCNE, decreased the expression of P27, and promoted the proliferation of TECs.

Enhancing the productivity and proliferation of CHO-K1 cells by oncoprotein YAP (Yes-associated protein).

CHO cells are extensively employed in biological drug industry to manufacture therapeutic proteins. Nevertheless, production of biopharmaceuticals faces obstacles such as limited growth and inadequate productivity. Employing host cell engineering techniques for CHO cells serves as a valuable approach to address the constraints encountered in biologics manufacturing. Despite advancements, most techniques focus on specific genes to address individual cellular challenges. The significance of YAP, transcriptional co-activator, cannot be overstated due to its involvement in regulating organ size and tumor formation. YAP's influence extends to various cellular processes and is regulated by kinase cascade in the Hippo pathway, which phosphorylates serine residues in specific LATS recognition motifs. Activation of YAP has been observed to impact both the size and quantity of cells. This research investigates the effects of YAP5SA on proliferation, apoptosis, and productivity in CHO-K1 cells. YAP5SA, with mutations in all five LATS-target sites, is selected for its heightened activity and resistance to repression through the Hippo-LATS1/2 kinase signaling pathway. Plasmid harboring YAP5SA was transfected into EPO-CHO and the influence of YAP5SA overexpression was investigated. According to our findings, transfection of EPO-CHO cells with YAP5SA exhibited a substantial enhancement in CHO cell productivity, resulting in a 3-fold increase in total protein and EPO, as well as a 1.5-fold increase in specific productivity. Additionally, it significantly contributes in augmenting viability, size, and proliferation. Overall, the findings of this study exemplify the potential of utilizing YAP5SA to impact particular cellular mechanisms, thereby presenting an avenue for customizing cells to fulfill production demands. KEY POINTS: • YAP5SA in CHO cells boosts growth, reduces apoptosis, and significantly improves productivity. • YAP5SA regulates genes involved in proliferation, survival, and mTOR activation. • YAP5SA increases productivity by improving cell cycle, c-MYC expression, and mTOR pathway.

CircROR1 upregulates CCNE1 expression to promote melanoma invasion and metastasis by recruiting KAT2A.

Circular RNAs (circRNAs) play important roles in cancer occurrence and progression. To explore and elucidate the clinical significance of specific circular RNA in melanoma and its potential molecular mechanism. CircROR1 expression in melanoma cells and tissues was confirmed by qRT-PCR and ISH. qRT-PCR and Western blotting were performed to measure the levels of CCNE1, KAT2A, MMP9 and TIMP2. MTT, Transwell and wound healing assays were performed to evaluate cell proliferation, invasion and metastasis. A xenograft mouse model was established to further verify the CircROR1/CCNE1 axis in vivo. RNA pull-down and RIP assays were performed to detect the direct interaction KAT2A and CircROR1. A ChIP assay was used to investigate the enrichment of H3K9ac acetylation in the CCNE1 promoter. CircROR1 was significantly upregulated in metastatic melanoma cells and tissues, promoting proliferation, invasion and metastasis in vitro and tumour growth in vivo. CircROR1 overexpression increased CCNE1 and MMP9 protein expression and decreased TIMP2 protein expression. Functional rescue assays demonstrated that CircROR1 played a role in promoting malignant progression through CCNE1. CircROR1 specifically bound to the KAT2A protein without affecting its expression. CircROR1 overexpression increased the level of H3K9ac modification in the CCNE1 promoter region by recruiting KAT2A, thus upregulating CCNE1 expression. CircROR1 upregulates CCNE1 expression through KAT2A-mediated histone acetylation. Our research confirms the critical role of CircROR1 in melanoma invasion and metastasis, and CircROR1 could serve as a potential therapeutic target for melanoma treatment.

Potential toxicity of graphene (oxide) quantum dots via directly covering the active site of anterior gradient homolog 2 protein.

Graphene quantum dots (GQDs) have attracted significant attention in biomedicine, while extensive investigations have revealed a reverse regarding the potential biotoxicity of GQDs. In order to supplementing the understanding of the toxicity profile of GQDs, this study employs a molecular dynamics (MD) simulation approach to systematically investigate the potential toxicity of both GQDs and Graphene Oxide Quantum Dots (GOQDs) on the Anterior Gradient Homolog 2 (AGR2) protein, a key protein capable of protecting the intestine. We construct two typical simulation systems, in which an AGR2 protein is encircled by either GQDs or GOQDs. The MD results demonstrate that both GQDs and GOQDs can directly make contact with and even cover the active site (specifically, the Cys81 amino acid) of the AGR2 protein. This suggests that GQDs and GOQDs have the capability to inhibit or interfere with the normal biological interaction of the AGR2 active site with its target protein. Thus, GQDs and GOQDs exhibit potential detrimental effects on the AGR2 protein. Detailed analyses reveal that GQDs adhere to the Cys81 residue due to van der Waals (vdW) interaction forces, whereas GOQDs attach to the Cys81 residue through a combination of vdW (primary) and Coulomb (secondary) interactions. Furthermore, GQDs aggregation typically adsorb onto the AGR2 active site, while GOQDs adsorb to the active site of AGR2 one by one. Consequently, these findings shed new light on the potential adverse impact of GQDs and GOQDs on the AGR2 protein via directly covering the active site of AGR2, providing valuable molecular insights for the toxicity profile of GQD nanomaterials.

Genome-wide CRISPR activation screen identifies JADE3 as an antiviral activator of NF-kB-dependent IFITM3 expression.

The innate immune system features a web of interacting pathways that require exquisite regulation. To identify novel nodes in this immune landscape, we conducted a gain-of-function, genome-wide CRISPR activation screen with influenza A virus. We identified both appreciated and novel antiviral genes, including Jade family PHD zinc finger 3 (JADE3) a protein involved in directing the histone acetyltransferase histone acetyltransferase binding to ORC1 complex to modify chromatin and regulate transcription. JADE3 is both necessary and sufficient to restrict influenza A virus infection. Our results suggest a distinct function for JADE3 as expression of the closely related paralogs JADE1 and JADE2 does not confer resistance to influenza A virus infection. JADE3 is required for both constitutive and inducible expression of the well-characterized antiviral gene interferon-induced transmembrane protein 3 (IFITM3). Furthermore, we find JADE3 activates the NF-kB signaling pathway, which is required for the promotion of IFITM3 expression by JADE3. Therefore, we propose JADE3 activates an antiviral genetic program involving NF-kB-dependent IFITM3 expression to restrict influenza A virus infection.

Human endogenous retrovirus type K encoded Np9 oncoprotein induces DNA damage response.

Human endogenous retrovirus sequences (HERVs) constitute up to 8% of the human genome, yet not all HERVs remain silent passengers within our genomes. Some HERVs, especially the HERV type K (HERV-K), have been found to be frequently transactivated in a variety of inflammatory diseases and human cancers. Np9, a 9-kDa HERV-K encoded protein, has been reported as an oncoprotein and found present in a variety of tumors and transformed cells. In the current study, we for the first time reported that ectopic expression of Np9 protein was able to induce DNA damage response from host cells especially through upregulation of γH2AX. Furthermore, we found that direct knockdown of Np9 by RNAi in Kaposi's Sarcoma-associated herpesvirus (KSHV) infected cells effectively reduced LANA expression, the viral major latent oncoprotein in vitro and in vivo, which may represent a novel strategy against virus-associated malignancies.

Histone lysine demethylase 4 family proteins maintain the transcriptional program and adrenergic cellular state of MYCN-amplified neuroblastoma.

Neuroblastoma with MYCN amplification (MNA) is a high-risk disease that has a poor survival rate. Neuroblastoma displays cellular heterogeneity, including more differentiated (adrenergic) and more primitive (mesenchymal) cellular states. Here, we demonstrate that MYCN oncoprotein promotes a cellular state switch in mesenchymal cells to an adrenergic state, accompanied by induction of histone lysine demethylase 4 family members (KDM4A-C) that act in concert to control the expression of MYCN and adrenergic core regulatory circulatory (CRC) transcription factors. Pharmacologic inhibition of KDM4 blocks expression of MYCN and the adrenergic CRC transcriptome with genome-wide induction of transcriptionally repressive H3K9me3, resulting in potent anticancer activity against neuroblastomas with MNA by inducing neuroblastic differentiation and apoptosis. Furthermore, a short-term KDM4 inhibition in combination with conventional, cytotoxic chemotherapy results in complete tumor responses of xenografts with MNA. Thus, KDM4 blockade may serve as a transformative strategy to target the adrenergic CRC dependencies in MNA neuroblastomas.

Targeted and Shallow Whole-Genome Sequencing Identifies Therapeutic Opportunities in p53abn Endometrial Cancers.

PURPOSE: Shallow whole-genome sequencing (sWGS) can detect copy-number (CN) aberrations. In high-grade serous ovarian cancer (HGSOC) sWGS identified CN signatures such as homologous recombination deficiency (HRD) to direct therapy. We applied sWGS with targeted sequencing to p53abn endometrial cancers to identify additional prognostic stratification and therapeutic opportunities. EXPERIMENTAL DESIGN: sWGS and targeted panel sequencing was performed on formalin-fixed, paraffin-embedded p53abn endometrial cancers. CN alterations, mutational data and CN signatures were derived, and associations to clinicopathologic and outcomes data were assessed. RESULTS: In 187 p53abn endometrial cancers, 5 distinct CN signatures were identified. Signature 5 was associated with BRCA1/2 CN loss with features similar to HGSOC HRD signature. Twenty-two percent of potential HRD cases were identified, 35 patients with signature 5, and 8 patients with BRCA1/2 somatic mutations. Signatures 3 and 4 were associated with a high ploidy state, and CCNE1, ERBB2, and MYC amplifications, with mutations in PIK3CA enriched in signature 3. We observed improved overall survival (OS) for patients with signature 2 and worse OS for signatures 1 and 3. Twenty-eight percent of patients had CCNE1 amplification and this subset was enriched with carcinosarcoma histotype. Thirty-four percent of patients, across all histotypes, had ERBB2 amplification and/or HER2 overexpression on IHC, which was associated with worse outcomes. Mutations in PPP2R1A (29%) and FBXW7 (16%) were among the top 5 most common mutations. CONCLUSIONS: sWGS and targeted sequencing identified therapeutic opportunities in 75% of patients with p53abn endometrial cancer. Further research is needed to determine the efficacy of treatments targeting these identified pathways within p53abn endometrial cancers.